Role of cytokinin in plant
development
Cytokinin – substance crucial
for sustain of cell proliferation
Cytokinins
Agrobacterium tumefaciens – IPT (isopentenyltransferase)
gene for cytokinin biosynthesis
Cytokinin biosynthesis
Takei et al., 2001
Isolation of Arabidopsis IPTs
Kakimoto 2004
Arabidopsis IPTs (8)
Differential expression patterns
Differently responsive promotors
(cytokinin, auxin, nitrate, combinations)
mutants, overexpression ???
CYTOKININ - Metabolism
Synthesis – IPT genes
Degradation – CK-oxidase
Conjugation
Cytokinin –
role in plant development
Auxin and cytokinin
Skoog et Miller, 1957
roots
shoots
Werner et al., 2001
Isolation of CK-oxidase (AtCKX)
AtCKXs overexpression in tobacco
Werner et al., 2001
Effect of AtCKX on tobacco root
Werner et al., 2001
Morphology of AtCKX tobacco plants
CK in shoot – positive regulator
CK in root – negative regulator
Werner et al., 2001
Cytokinin – root meristem development
Dello Ooio et al., 2007
Cytokinin - lateral root organogenesis
Laplaze et al., 2007
Cytokinin effect is stage specific
Laplaze et al., 2007
Cytokinin
–signal perception and transduction
Kakimoto et al., 1996
Isolation of CK independent (cki1) mutant
Identification of CKI1 gene
Kakimoto et al., 1996
Verification - 35S::CKI1 transgene
+CK
-CK
Kakimoto et al., 1996
Inoue et al., 2001
Next strike
- CK response
mutant (cre1)
Wt
cre1
pGTV pGTV-CRE1
His kinase transduction pathway
Piece of genius - complementation
Hog1 kinase hyperactivation
Inoue et al., 2001
Cytokinin receptors – what else?
3 CRE1 homologous proteins (AHKs)
multiple mutant phenotypes – additive
(not lethal)
What does CKI1?
His kinase transduction pathway
His kinase pathway
components
in Arabidopsis
Response Regulators in Arabidopsis
Brandstatter and Kieber, 1999
CK responsive genes –
ARR type A
Response regulator ARR type Anegative
regulators of cytokinin signalling
To et al., 2004
Phenotypes of arr type B positive regulator
of CK signalling
wt
35S::ARR1
arr1-1
Sakai et al., 2001
Hutchison et al.,2006
AHPs mediate transfer of cytokinin signal between
cytoplasm and nucleus
Hwang
Games with protoplasts
Opposite effects of two classes
of ARRs on CK signalling
AHPs – signal to nucleus
Hextuple of type A ARRs
confirms role as negative
regulators
e.g. Root elongation
assay
Transcritpion
Cell division
Shoot formation
Delayed senescence
Vascular develppment
Cytokinin – root meristem development
Dello Ioio et al., 2007
Blilou et al.,2005
Auxin related mutants with short
meristem
Auxin transport mutantsMS cytokinin
Cytokinin modulates auxin transport
MS BA
PIN2:GFP (48 h)
PIN7:GFP (24h)PIN3:GFP (24h)
PIN1:GFP (24 h)
MS BA
MS BA MS BA
*
* *
* * *
*
*
*
* * * * *
0
50
100
150
200
250
300
PIN1 PIN2 PIN3 PIN7
Relativeexpression(%)
MS
1h
2h
3h
4h
5h
6h
Q-RT PCR
Ruzicka et al., 2009
MS BA
* * *
Cytokinin reduces auxin efflux in tobacco BY2 cells
PIN1:RFP
Ruzicka et al., 2009
CK receptor CK response regulator AUX/IAA negative regulator
CK – auxin signalling pathways interaction
to modulate auxin transport
Expression of IAA3/SHY2
is CK regulated
Dello Ioio et al., 2008
Model of cytokinin and auxin interaction in root meristem
Dello Ioio et al., 2008
Cytokinin signalling regulates
vascular development
Mahonen et al., 2006
ahp6 suppress wol insensitivity
to cytokinin
AHP6 codes for
histidine phosphotransfer protein 6
AHP6 is negative regulator
of cytokinin signalling